Instrument and method for the automated thermal treatment of liquid samples

ABSTRACT

An instrument and a method for the automated thermal treatment of liquid samples are disclosed. An inter-distance between a temperature-controlled receptacle for loading with a plurality of vessels for containing the samples and end portions of optical fibers can be varied, wherein the receptacle is configured to form a thermal communication with the loaded vessels and wherein the optical fibers have first and second end portions. The first end portion and the second end portion of each optical fiber is fixed with respect to each other for transmitting light, wherein the variation of the inter-distance allows the vessels to be loaded to or unloaded from the receptacle and to enable detection of light from the samples contained in the one or more receptacle-loaded vessels.

The present application is a continuation of application Ser. No.13/295,504 filed on Nov. 14, 2011 which claims priority to EPApplication Number 10191156.8 filed Nov. 15, 2010, the priority of bothapplications is claimed hereby and the disclosures of both applicationsare hereby incorporated by reference.

TECHNICAL FIELD

The embodiments of the present invention are in the field of biochemicalresearch, biochemical routine analytics, clinical diagnostics andclinical research and more particularly concerns an instrument andmethod for the automated thermal treatment and, e.g., fluorescencedetection of liquid samples.

BACKGROUND

In these days, nucleic acids (DNA=deoxyribonucleic acid, RNA=ribonucleicacid) are subject to various analyses and assays in the above-describedtechnical field. In order to detect small amounts, the well-knownpolymerase chain reaction (PCR) can be used to replicate the targetnucleic acid sequence to an amount which is detectable. Theamplification of nucleic acids using the polymerase chain reaction hasbeen extensively described in the patent literature, for instance, inU.S. Pat. Nos. 4,683,303, 4,683,195, 4,800,159 and 4,965,188. Generally,in the polymerase chain reaction, samples containing reaction mixturesof specific reagents and nucleic acids are repeatedly put through asequence of amplification steps. Each sequence includes melting thedouble-stranded nucleic acids to obtain denaturated singlepolynucleotide strands, annealing short primers to the strands andextending those primers to synthesize new polynucleotide strands alongthe denaturated strands to make new copies of double-stranded nucleicacids. Due to the fact that reaction conditions strongly vary withtemperatures, the samples are put through a series of temperatureexcursions in which predetermined temperatures are kept constant forspecific time intervals (“thermo-cycling”). The temperature of thesamples typically is raised to around 90° C. for melting the nucleicacids and lowered to a temperature in the range of from 40° C. to 70° C.for annealing and primer extension along the polynucleotide strands.

It is also known to detect the PCR reaction products during progress ofthe polymerase chain reaction (“real-time PCR”) to detect the presenceor absence of a target nucleic acid sequence (or analyte) and/or toquantify the original amount of target nucleic acid which was present inthe sample. In daily routine, commercially available instruments arebeing used for performing the PCR and detecting the reaction productsobtained by means of fluorescence.

SUMMARY

In one embodiment, an instrument for the automated thermal treatment ofliquid samples is disclosed. The instrument may comprise atemperature-controlled receptacle for loading with a plurality ofvessels for containing the samples, the receptacle being configured toform a thermal communication with the loaded vessels. A detection moduleequipped with a detection arrangement may be provided with one or moredetectors for detecting light emitted from the samples and a couplingarrangement provided with a plurality of optical fibers for transmittingthe emitted light to the detection arrangement, wherein the opticalfibers have first and second end portions, and the first end portion andthe second end portion of each optical fiber are fixed with respect toeach other. A moving mechanism for moving at least one of the couplingarrangement and the receptacle in a manner to vary an inter-distancebetween the coupling arrangement and the receptacle is provided so as toallow the vessels to be loaded to or unloaded from the receptacle and toallow detection of light from samples contained in the one or morereceptacle-loaded vessels.

In another embodiment, a method for the automated thermal treatment ofliquid samples is disclosed. The method may comprise varying aninter-distance between a temperature-controlled receptacle for loadingwith a plurality of vessels for containing the samples and end portionsof optical fibers, wherein the receptacle is configured to form athermal communication with the loaded vessels and wherein the opticalfibers have first and second end portions. The first end portion and thesecond end portion of each optical fiber are fixed with respect to eachother for transmitting light. The variation of the inter-distance allowsthe vessels to be loaded to or unloaded from the receptacle and to allowdetection of light from the samples contained in the one or morereceptacle-loaded vessels.

These and other embodiments are disclosed hereafter in the detaileddescription as well as in the accompanying drawings.

BRIEF DESCRIPTION OF THE DRAWINGS

The accompanying drawings, which are incorporated in and constitute apart of the specification, illustrate certain embodiments of theinvention, and in which:

FIG. 1 is a schematic illustration of various modules of exemplaryinstruments for the automated thermal treating of samples according tothe invention;

FIG. 2 is a partly sectioned perspective view of an exemplary instrumentthe modules of which are schematically illustrated in FIG. 1;

FIG. 3 is a sectional view of the instrument of FIG. 2;

FIG. 4 illustrates an enlarged detail of the instrument of FIGS. 2 to 3;

FIG. 5 illustrates another enlarged detail of the instrument of FIGS. 2to 3 according to circle A of FIG. 4;

FIG. 6 illustrates another enlarged detail of the instrument of FIGS. 2to 5;

FIGS. 7A-7B are sectional partial views of the instrument of FIGS. 2 to6 illustrating a vertical movement of the detection module;

FIG. 8 illustrates a fixing element for fixing the second end portionsof the optical fibers of the instrument of FIGS. 2 to 7;

FIGS. 9A-9F are perspective views illustrating a tray for moving thethermal module of the instrument of FIGS. 2 to 8;

FIG. 10 is a partly sectioned perspective view of another exemplaryinstrument the modules of which are schematically illustrated in FIG. 1;

FIG. 11 illustrates a one-to-one connection between the first and secondend portions of the optical fibers of the instruments of FIGS. 2 to 10;

FIG. 12 illustrates a device for mapping the first and second ends ofthe optical fibers of the instruments of FIGS. 2 to 10;

FIG. 13 is a sectional view illustrating a clamping mechanism forclamping the receptacle of the instruments of FIGS. 2 to 10;

FIG. 14 illustrates an enlarged detail of FIG. 13; and

FIG. 15 is a perspective view illustrating the clamp of FIGS. 13 and 14.

REFERENCE LIST

-   -   1 Instrument    -   2 Thermal module    -   3 Multi-well plate    -   4 Well    -   5 Sample    -   6 Detection module    -   7 Excitation arrangement    -   8 Light source    -   9 Excitation light    -   10 Detection arrangement    -   11 Detector    -   12 Coupling arrangement    -   13 Excitation fiber    -   14 Emission fiber    -   15 First end portion    -   16 First fixing element    -   17 Second end portion    -   18 Second fixing element    -   19 Third fixing element    -   20 Excitation optics    -   21 Emission optics    -   22 Excitation filter    -   23 Emission filter    -   24 Emitted light    -   25 Controller    -   26 Electric lines    -   27 Thermal block    -   28 Receptacle    -   29 Heat exchanger    -   30 Rib    -   31 Main base    -   32 Upper receptacle face    -   33 Recess    -   34 Sealing cover    -   35 Upper plate face    -   36 Lower plate face    -   37 Contact face    -   38 Chassis    -   39 Vertical plate    -   40 Upper horizontal plate    -   41 Lower horizontal plate    -   42 First through-hole    -   43 First opening    -   44 Second opening    -   45 Second through-hole    -   46 Third through-hole    -   47 First casing    -   48 Dichroic mirror    -   49 Second casing    -   50 Filter wheel    -   51 Electric motor    -   52 Moving mechanism    -   53 Rail    -   54 Free space    -   55 Thermoelectric device    -   56 Cylindrical portion    -   57 Disk-like portion    -   58 Projection    -   59 Orifice    -   60 First end face    -   61 Second end face    -   62 Hollow    -   63 Adhesive material    -   64 Cover heater    -   65 Heating element    -   66 Heating plate    -   67 Plate hole    -   68 Cavity    -   69 Upper plate face    -   70 Element hole    -   71 Contact region    -   72 Planar portion    -   73 Lower planar portion face    -   74 Upper heating element face    -   75 Upper horizontal plate face    -   76 Core    -   77 Coating    -   78 Cover heater hole    -   79 Mapping device    -   80 Base    -   81 Light generating element    -   82 Control panel    -   83 First turn-switch    -   84 Second turn-switch    -   85 Device controller    -   86 Electric line    -   87 Upper base face    -   88 Contact area    -   89 Base    -   90 Instrument casing    -   91 Lever    -   92 Fulcrum    -   93 Upper lever portion    -   94 Lower lever portion    -   95 Upper end    -   96 Connecting rod    -   97 Turning knuckle    -   98 First arm    -   99 Second arm    -   100 Guiding rod    -   101 Coil spring    -   102 Tray    -   103 Supporting base    -   104 Beam    -   105 Front cover    -   106 Recessed grip    -   107 Spring catch    -   108 Opening/closing device    -   109 Central axis    -   110 Lower end    -   112 Projection    -   113 Guiding rail    -   114 Resting portion    -   115 Data storage    -   116 Reference channel    -   117 Reference channel fiber    -   118 Reference channel detector    -   119 Flange    -   120 Screw    -   121 Mounting hole    -   122 Interface layer    -   123 Clamp    -   124 Isolation block    -   125 Base plate    -   126 Arm    -   127 Middle portion    -   128 Gripping portion    -   129 Clamping mechanism    -   130 Gripping recess

DETAILED DESCRIPTION

According to a first aspect, a new instrument for the automated thermaltreatment of liquid samples is disclosed. In some embodiments, theinstrument is being used for the execution of the PCR, in particularreal-time PCR. Specifically, in some embodiments, the instrument isbeing used for PCR with hybridization probes, PCR with hydrolysisprobes, PCR with interchelator dyes, real-time PCR with correspondingprobes, various isothermal amplification methods with correspondingfluorescence reporters and melting analysis of DNA. Typical analyses arethe detection of the presence/absence and optionally concentration ofpathogens such as virus or bacteria in a sample, genotyping, measuringexpression profiles, and many others.

In some embodiments, the instrument comprises a temperature-controlledreceptacle for receiving a plurality of vessels for containing theliquid samples. The receptacle is being configured to form a thermalcommunication with the loaded vessels so that samples contained thereinare in thermal communication with the receptacle to be heated or cooledaccording to the specific demands of the user.

In some embodiments, the instrument comprises a detection moduleequipped with a detection arrangement provided with one or moredetectors for detecting light emitted from the liquid samples and acoupling arrangement provided with a plurality of optical fibers fortransmitting emitted light to the detection arrangement, wherein saidoptical fibers have first and second end portions, and wherein, in someembodiments, the first end portion and the second end portion of eachoptical fiber are being fixed with respect to each other.

In some embodiments, the detection module further comprises anexcitation arrangement provided with one or more light sources forgenerating excitation light. In some embodiments, the one or moredetectors are configured for detecting light emitted from the liquidsamples in response to the excitation light.

In some embodiments, the instrument comprises a moving mechanism suchas, but not limited to, an automated moving mechanism for moving thecoupling arrangement and/or the receptacle in a manner to vary aninter-distance between the coupling arrangement and the receptacle so asto allow the vessels to be loaded to or unloaded from the receptacle sothat the vessels, i.e., samples contained therein can be brought in andout of thermal communication with the receptacle and to allow thedetection of light from samples contained in the one or morereceptacle-loaded vessels. In some embodiments, the moving mechanismincludes a controllable drive such as, but not limited to, an electricmotor or a hydraulic or pneumatic actuator operatively coupled to aguiding mechanism such as, but not limited to, a rack-and-pinionmechanism for automatically moving the coupling arrangement and/or thereceptacle so as vary an inter-distance between the coupling arrangementand the receptacle.

In some embodiments, the moving mechanism is configured to move thecoupling arrangement while the receptacle is being kept stationary. Insome embodiments, the moving module is configured to move the detectionmodule. In some embodiments, the moving mechanism is configured to movethe coupling arrangement while the detection arrangement is being keptstationary.

In some embodiments, the optical fibers include first fibers, in thefollowing denoted as “emission fibers” for transmitting the emittedlight to the detection arrangement and second fibers different from thefirst fibers, in the following denoted as “excitation fibers” fortransmitting excitation light to the samples. In some embodiments, thefirst end portions of the excitation and emission fibers are being fixedby at least one plate-like fixing element, wherein the second endportions of the excitation and emission fibers are being fixed by atleast one another plate-like fixing element. Specifically, in someembodiments, the first end portions of the excitation and emissionfibers are being fixed by one first plate-like fixing element, whereinthe second end portions of the excitation fibers are being fixed by onesecond plate-like fixing element and the second end portions of theemission fibers are being fixed by one third plate-like fixing element.

In some embodiments, the coupling arrangement is being equipped with acover heater for heating a sealing cover placed over a multi-well platehaving a plurality of wells for containing the samples. In someembodiments, the cover heater includes a heated plate-like heatingmember configured to be brought in physical contact with the sealingcover, wherein the heating member is being equipped with a plurality ofopenings accommodating first end portions of the optical fibers. In someembodiments, the optical fibers are being thermally isolated from theheating member. In some embodiments, the openings are configured to formcavities in case the heating member contacts the multi-well plate,wherein the cavities are being adapted to optically shield the wellsfrom each other. In some embodiments, the openings are configured toform closed cavities in case the heating member contacts the multi-wellplate. In some embodiments, the heating member is configured to exertmechanical pressure on the multi-well plate so as to press the wellsinto recesses of the receptacle.

In some embodiments, the instrument includes a controller set up tocontrol the instrument for the thermal treatment of samples. In someembodiments, the controller is configured as programmable logiccontroller running a machine-readable program provided with instructionsto perform operations for thermally treating the liquid samples. Statedmore particularly, in some embodiments, the controller is electricallyconnected to the components requiring control. In some embodiments, thecontroller is set up to perform a step of varying an inter-distancebetween the temperature-controlled receptacle and the first end portionsof the optical fibers. In some embodiments, the controller is set up toperform a step of moving the optical fibers with respect to thereceptacle while keeping the receptacle stationary. In some embodiments,the controller is set up to perform a step of commonly moving theoptical fibers and the detection arrangement with respect to thereceptacle while keeping the receptacle stationary. In some embodiments,the controller is designed to execute the fluorescence detection of thesamples, including control of at least one light source, position offilter wheels, operation of one or more detectors and data processing.

In some embodiments, the instrument includes or may access a volatile ornon-volatile data storage storing an end-to-end relationship (mapping)between the first end portion and the second end portion of individualoptical fibers so that the second end portion of each optical fiber canbe assigned to the first end portion thereof in a one-to-onerelationship. The mapping in particular concerns the position of thefirst end portion in terms of from which vessel light is received, thatis to say, each optical fiber is positioned to receive light from aspecific sample vessel located beneath. The second end portions have astochastic lateral position and light from the second ends is receivedby a laterally-resolving detector. Due to the end-to-end mapping, it isdetermined in which lateral position light from a particular vessel isreceived. Accordingly, a signal measured can be related by theend-to-end mapping to a specific sample vessel.

In some embodiments, the coupling arrangement is being provided with thedata storage storing the end-to-end relationship (mapping). In someembodiments, the controller relates to measured fluorescence data froman optical sensor array to respective vessels—based on the mapping datain the data storage. Non-volatile data storages are preferred becausedata can be stored right after the mapping process and the non-volatiledata storage can be fixed to the coupling arrangement so that a mix-upand unintended data manipulations can mostly be prevented. Examples forthe non-volatile data storage are compact disc (CD), USB-stick, EEPROM,flash memory. The instrument can also be supplied with the end-to-endrelationship (mapping) via the internet. Access to the internet can beprovided by a LAN or WLAN or UMTS-connection or any other wired orwireless connection technique.

According to a second aspect, a new method for the automated thermaltreatment and, e.g., fluorescence detection of liquid samples isdisclosed. The method can, e.g., be implemented in the above-describedinstrument for thermally treating liquid samples.

In some embodiments, the method comprises a step of varying aninter-distance between a temperature-controlled receptacle for loadingwith a plurality of vessels for containing the samples and end portionsof optical fibers. Specifically, the receptacle is being configured toform a thermal communication when the vessels are loaded on thereceptacle. Otherwise, the optical fibers have first and second endportions, wherein the first end portion and the second end portion ofeach optical fiber is being fixed with respect to each other fortransmitting light emitted from the samples. By varying aninter-distance between the receptacle and the first end portions of theoptical fibers, the vessels can be loaded to or unloaded from thereceptacle to be brought in and out of thermal communication with thereceptacle and light can be detected from the samples contained in theone or more receptacle-loaded vessels.

In some embodiments, the first end portions of the optical fibers aremoved with respect to the receptacle while keeping the receptaclestationary. In some embodiments, the end portions of the optical fibersand the detection arrangement are commonly moved with respect to thereceptacle while keeping the receptacle stationary.

According to a third aspect, a new device for determining a mappingbetween end portions of optical fibers in an instrument for thermallytreating liquid samples is disclosed.

In some embodiments, the mapping device comprises a plate-like baseconfigured to be put on a temperature-controlled receptacle forreceiving a plurality of vessels for containing the liquid samples inthermal communication therewith. In some embodiments, the base is beingprovided with a plurality of light generating elements for generatinglight, wherein the light generating elements are being arranged in amanner to be optically coupled with a plurality of optical fibersconfigured for transmitting light emitted from the samples to at leastone detector. In some embodiments, the mapping device comprises acontroller set up for selectively supplying electric current to thelight generating elements.

According to a fourth aspect, a new method for determining a mappingbetween end portions of optical fibers in an instrument for thermallytreating liquid samples is disclosed.

In some embodiments, the method comprises a step of putting a plate-likebase on a temperature-controlled receptacle for receiving a plurality ofvessels for containing the samples in thermal communication therewith.In some embodiments, the method comprises a step of selectivelysupplying electric current to a plurality of light generating elementsfor generating light. In some embodiments, the method comprises a stepof optically coupling the light into optical fibers configured fortransmitting light emitted by the samples. In some embodiments, themethod comprises a step of detecting the light exiting the opticalfibers by at least one detector.

The above-described embodiments of the various aspects of the inventionmay be used alone or in any combination thereof without departing fromthe scope of the invention.

Various illustrated embodiments according to the present invention willbe described in detail below with reference to the accompanyingdrawings, where like designations denote like or similar elements. Firstreferring to FIGS. 1 through 10 embodiments of exemplary instruments forthe automated thermal treating of liquid samples generally referred toat reference numeral 1 are explained. FIG. 1 illustrates various modulesof the exemplary instruments depicted in FIGS. 2 to 10. Specifically,FIGS. 2 to 9 refer to a first exemplary instrument 1; FIG. 10 refers toa second exemplary instrument 1 only differing from the first one in theexcitation and detection arrangements. In some embodiments, theinstrument 1 is configured as a thermo-cycler for thermally cyclingreaction mixtures of nucleic acids and one or more reagents through aseries of temperature excursions and optically detecting the reactionproducts obtained. In some embodiments, the instrument 1 can be used toperform the PCR, in particular real-time PCR, or any other reaction ofthe nucleic acid amplification type. In some embodiments, the instrument1 can be used for the optical on-line detection of reaction products. Insome embodiments, the instrument 1 can be used for the isothermaltreatment or execution of melting curves.

With particular reference to FIG. 1, in some embodiments, the instrument1 includes various modules as detailed in the following descriptionwhich are functional and (optionally) structural entities for treatingliquid samples. Specifically, in some embodiments, the instrument 1includes a thermal module 2 which can be brought in thermalcommunication with a multi-well plate 3 provided with plural vessels,cavities or wells 4 for receiving liquid samples 5. In some embodiments,the thermal module 2 can be heated or cooled according to pre-definedtemperature profiles so as to transfer heat in controlled manner to/fromthe samples 5.

With continued reference to FIG. 1, in some embodiments, a detectionmodule 6 can be used to detect light so as to identify reaction productswhich, in some embodiments, can be obtained as a result of a polymerasechain reaction of the samples 5. In some embodiments, the instrument 1can be used for the optical on-line detection of the reaction productsduring progress of the amplification reactions.

As indicated by the double arrows, in some embodiments, the detectionmodule 6 can at least vertically be moved in controlled manner relativeto the thermal module 2 by means of a moving mechanism 52 (not furtherdetailed in FIG. 1). In some embodiments, the moving mechanism 52 is anautomated moving mechanism. The moving mechanism 52 can, e.g., beconfigured as driven rack and pinion mechanism or any other mechanismenabling at least a vertical movement of the detection module 6.Specifically, the moving mechanism 52 can, e.g., include a(controllable) driver such as, but not limited to, an electric motor orhydraulic actuator for automatically moving the detection module 6.Since those of skill in the art are aware of the specific configurationof such moving mechanism, it is not necessary to elucidate it furtherherein. Using the moving mechanism 52, in some embodiments, thedetection module 6 can selectively be moved into a lowered first oroperative position adapted for optically detecting reaction productsobtained from the samples 5 or in a raised second or inoperative orloading/unloading position adapted for loading or unloading theinstrument 1 with the multi-well plate 3.

With continued reference to FIG. 1, in some embodiments, the detectionmodule 6 includes an excitation arrangement 7 provided with at least onelight source 8 for generating excitation light 9 adapted to excite theemission of light 24 (e.g. fluorescence light), in the following denotedas “emitted light”, by the samples 5. As illustrated, in someembodiments, the detection module 6 further includes a detectionarrangement 10 provided with at least one detector 11 to opticallydetect the emitted light 24. In some embodiments, the detection module 6further includes a coupling arrangement generally referred to atreference numeral 12 for optically coupling each of the excitationarrangement 7 and the detection arrangement 10 to the wells 4. Statedmore particularly, in some embodiments, the coupling arrangement 12includes plural first optical fibers 13, in the following denoted as“excitation fibers”, for transmitting the excitation light 9 from theexcitation arrangement 7 to the wells 4 and plural second optical fibers14, in the following denoted as “emission fibers”, for transmitting theemitted light 24 from the wells 4 to the detection arrangement 10. Insome embodiments, each well 4 of the multi-well plate 3 is related to anindividual pair of one excitation fiber 13 and one emission fiber 14.

As further illustrated in FIG. 1, in some embodiments, well-sided firstend portions 15 of the excitation fibers 13 are fixed with respect toeach other by means of a first fixing element 16, while second endportions 17 of the excitation fibers 13 opposite to the first endportions 15 thereof are fixed with respect to each other by a secondfixing element 18. Otherwise, in some embodiments, well-sided first endportions 15 of the emission fibers 14 are fixed with respect to eachother by means of the first fixing element 16, while second end portions17 of the emission fibers 14 opposite to the first end portions 15thereof are fixed with respect to each other by a third fixing element19. Specifically, in some embodiments, the excitation light 9 can becoupled into the excitation fibers 13 at second end faces 61 and becoupled out of the excitation fibers 13 at first end faces 60 thereof.Otherwise, in some embodiments, the emitted light 24 can be coupled intothe emission fibers 14 at first end faces 60 and be coupled out of theemission fibers 14 at second end faces 61.

With continued reference to FIG. 1, in some embodiments, an excitationoptics generally referred to at reference numeral 20 is used tooptically couple the excitation light 9 into the excitation fibers 13 atthe second end faces 61. Specifically, in some embodiments, one or moreexcitation filters 22 are used for filtering one or more specificwavelengths or one or more ranges of wavelengths before the excitationlight 9 is coupled into the excitation fibers 13. Otherwise, in someembodiments, in case the detection module 6 is in operative position,the first end faces 60 of the excitation fibers 13 are arranged in sucha manner that the excitation light 9 is directed into the wells 4 toexcite the emitted light 24 by the samples 5.

With continued reference to FIG. 1, in some embodiments, in case thedetection module 6 is in operative position, the first end faces 60 ofthe emission fibers 14 are arranged in such a manner that the emittedlight 24 can be coupled into the emission fibers 14. In someembodiments, an emission optics generally referred to at referencenumeral 21 is used to optically couple the emitted light 24 leaving theemission fibers 14 at the second end faces 61 to the detector 11. Insome embodiments, one or more emission filters 23 are used for filteringone or more wavelengths or one or more ranges of wavelengths from theemitted light 24 before the emitted light 24 hits the detector 11.

In the instrument 1, in some embodiments, one pair of optical fibers 13,14 is used as optical reference channel for performing referencemeasurements. Instead of a normal sample one or a set of referencesamples can be contained in a well 4 associated with the optical fibers13, 14. In some embodiments, the reference samples are made offluorescent glass or crystal such as, but not limited to, terbium glassor ruby.

With continued reference to FIG. 1, in some embodiments, the instrument1 has an implemented (dedicated) reference channel 116 configured tomonitor brightness and intensity of the light source 8 generating theexcitation light 9. The signal of the reference channel 116 can be usedto survey the light source 8 and/or to control the intensity of theexcitation light 9 so as to have a constant intensity, e.g., by means ofa feedback regulation loop. Alternatively the signal of the referencechannel 116 can be used to normalize the measured sample fluorescencedata. The reference channel 116 consists of one reference channel fiber117 configured to direct light generated by the light source 8 to areference channel detector 118 for measuring the intensity of lighttransmitted through the reference channel fiber 117. In someembodiments, the reference channel detector 118 is being connected to anelectronic amplifier (not illustrated) and an analogue digital converter(not illustrated).

In some embodiments, the controller 25 for controlling the automatedthermal treating of the samples 5 is configured as micro-controllerrunning a computer-readable program provided with instructions toperform operations in accordance with a pre-defined sequence of steps.Specifically, the controller 25 receives information from the variouscomponents of the instrument 1, especially from the detector 11, andgenerates and transmits corresponding control signals to the componentswhich require control such as the moving mechanism 52 for verticallymoving the detection module 6, the light source 8 and the thermal module2. As schematically illustrated in FIG. 1, in some embodiments, electriclines 26 are used for transmitting the electric signals.

With particular reference to FIGS. 2 to 10, exemplary embodiments of theinstrument 1 as schematically illustrated in FIG. 1 now are explained.Specifically, FIGS. 2 to 9 refer to a first variant of the exemplaryinstrument 1, while FIG. 10 refers to a second variant thereof.Accordingly, in some embodiments, the thermal module 2 includes atemperature-controlled (thermal) block 27 made of material having goodthermal conductivity such as metallic material. As illustrated in FIG.5, in some embodiments, the thermal block 27 is provided with one ormore thermoelectric devices 55 which, in some embodiments, utilize thePeltier effect. Connected to a DC power source (not illustrated), eachof the thermoelectric devices 55 functions as heat pump for producing orabsorbing heat depending upon the direction of the electric currentapplied. In some alternative embodiments, the thermoelectric devices 55are replaced by other heating devices such as resistive heaters whichcan be heated based on Ohmic heating in combination with cooling meanssuch as a fan for cooling the thermal block 27 with air. In somealternative embodiments, the thermal block 27 is provided with channelswhich can be flown-through by liquids having different temperatures.Generally other heating/cooling means as known from the prior art may beused.

In some embodiments, on an upper side thereof, the thermal block 27 isintegrally formed with a plate-like receptacle 28 adapted for holdingthe multi-well plate 3 in thermal communication with the thermal block27 and, in some embodiments, is made of material having good thermalconductivity so as to enable heat transfer to/from the samples 5contained in the wells 4. In some embodiments, the thermal module 2includes a heat exchanger 29 thermally coupled to the thermal block 27on a lower side thereof. Specifically, in some embodiments, the heatexchanger 29 is provided with plural plate-like ribs 30 seriallyarranged with respect to each other keeping a small inter-distance toenable effective heat transfer to the ambient.

In some embodiments, the thermoelectric devices 55 of the thermal module2 can be supplied with electric current to heat or cool the receptacle28 so as to change and hold various temperatures of the samples 5 for apredetermined amount of time under control of the controller 25.Specifically, in some embodiments, the controller 25 can transmitcontrol signals to the thermoelectric devices 55 to regulate the desiredtemperature of the receptacle 28 which, in some embodiments, is variedin response to the input of a temperature sensor (not illustrated) forsensing the temperature of the receptacle 28 and/or the samples 5.

As illustrated in FIG. 5, in some embodiments, an upper face 32 of thereceptacle 28, in the following denoted as “upper receptacle face” isprovided with plural recesses 33, the inner profiles of which areconform in shape with the outer contours of the wells 4 at least intheir lower parts so that the multi-well plate 3 can be placed upon thereceptacle 28 in a position where the wells 4 rest inside the recesses33. Accordingly, due to an at least partially close fit, good thermalcommunication between the receptacle 28 and the wells 4 can be obtainedresulting in a highly efficient transfer of heat between the receptacle28 and the wells 4. Otherwise, in some embodiments, a lower face 36 ofthe multi-well plate 3, in the following denoted as “lower plate face”is distanced from the upper receptacle face 32 in regions in-between thewells 4 to improve the thermal transfer between the receptacle 28 andthe wells 4.

In some embodiments, the multi-well plate 3 comprises a main base 31having an upper face 35, in the following denoted as “upper plate face”,which is provided with a rectangular array of wells 4 for receiving thesamples 5. The array may, e.g., include 8×12 wells (96 wells), 6×10wells (60 wells), 16×24 wells (384 wells), or any other number andarrangement that would be compatible with the automated instrument 1 forthermally treating the samples 5. The footprint of the multi-well plate3 may, e.g., be about 127 mm in length and about 85 mm in width, whilethose of skill in the art will recognize that the multi-well plate 3 canbe formed in dimensions other than those specified herein. In someembodiments, the multi-well plate 3 consists of plastic material such asbut not limited to polypropylene, polystyrene and polyethylene. In someembodiments, the multi-well plate 3 is intended for single use only sothat it can be filled with samples 5 for a single experiment and isthereafter discarded.

In some embodiments, a transparent sealing cover 34 is fixed to theupper plate face 35 at planar contact regions 71 thereof locatedin-between the wells 4 by adhesion or thermal sealing. Specifically, thetransparent sealing cover 34 air-tightly seals the open-top wells 4 inorder to prevent evaporation of the samples 5 and to shield the samples5 from external influences such as cross-contamination. In someembodiments, the transparent sealing cover 34 is made of an opticallytransparent material such as a clear film exhibiting low fluorescencewhen exposed to the excitation light 9. In some embodiments, thetransparent sealing cover 34 is made of one or more polymers selectedfrom the group consisting of polystyrene, polyethylene and polyester. Insome embodiments, the transparent sealing cover 34 is a multi-layeredfilm, e.g., consisting of one layer of polypropylene and one layer ofpolyester. In some embodiments, the transparent sealing cover 34comprises one or more compliant coatings and/or one or more adhesivessuch as a pressure sensitive adhesive or a hot melt adhesive for fixingthe transparent sealing cover 34 to the upper plate face 35. Thetransparent sealing cover 34 allows for an optical detection of theemitted light 24, e.g., during progress of the polymerase chain reactionso as to enable an optical on-line detection of the reaction productsobtained. The transparent sealing cover 34 thus allows the excitationlight 9 to be transmitted to the wells 4 and the emitted light 24 to betransmitted back to the one or more detectors 11. In some embodiments,the sealing cover 34 is applied to the multi-well plate 3 after thesamples have been filled into the wells 4 and before the multi-wellplate 3 is loaded in the instrument 1.

With particular reference to FIG. 2, in some embodiments, the couplingarrangement 12 comprises a rigid chassis 38 including four verticalplates 39, a upper horizontal plate 40 and two spaced-apart lowerhorizontal plates 41 assembled by conventional fixation means such asscrews, bolts or welded connections. In some embodiments, the firstfixing element 16 for fixing the first end portions 15 of both theexcitation and emission fibers 13, 14 is a rectangular solid plate fixedto the lower horizontal plates 41 so as to be integrally formed with thechassis 38. In some embodiments, each of the second and third fixingelements 18, 19 for fixing the second end portions 17 of the emissionand excitation fibers 13, 14, respectively, comprises a cylindricalportion 56 and a disk-like planar portion 57. As illustrated, in someembodiments, the cylindrical portions 56 are inserted into first andsecond openings 43, 44, respectively, formed by the upper horizontalplate 40 and are fixed therein like plugs. Fixed to the upper horizontalplate 40, the second and third fixing elements 18, 19 are integrallyformed with the chassis 38.

As, e.g., illustrated in FIG. 5, in some embodiments, the first fixingelement 16 is provided with an array of first through-holes 42. In someembodiments, the number and arrangement of the first through-holes 42correspond to the number and arrangement of the wells 4 wherein thefirst through-holes 42 can, e.g., be located right above the wells 4.With continued reference to FIG. 5, in some embodiments, each firstthrough-hole 42 accommodates the first end portions 15 of a pair of oneexcitation fiber 13 and one emission fiber 14 allowing the excitationlight 9 transmitted by the excitation fibers 13 to be directed into thewells 4 for irradiating the samples 5. Otherwise, the emitted light 24can be received by the emission fibers 14. In some alternativeembodiments, two first through-holes 42 are provided for each well 4,one for accommodating the first end portion 15 of the excitation fiber13 and the other one for accommodating the first end portion 15 of theemission fiber 14, so that each first end portion 15 is accommodated ina separate first through-hole 42. In some embodiments, the two firstthrough-holes 42 related to one well 4 are spaced apart to have adistance from each other in a range of from 0.1 to 2 mm. Those of skillin the art will recognize that inter-distances other than thosespecified herein can be envisaged according to the specific demands ofthe user.

With particular reference to FIG. 8 illustrating one of the second orthird fixing elements 18, 19, in some embodiments, the second fixingelement 18 is provided with one second through-hole 45 and the thirdfixing element 19 is provided with one third through-hole 46, whereinthe second through-hole 45 accommodates all second end portions 17 ofthe excitation fibers 13 and the third through-hole 46 accommodates allsecond end portions 17 of the emission fibers 14. Specifically, whileFIG. 8 depicts the second fixing element 18, the third fixing element 19is similar in construction as indicated by the reference signs inbrackets related to the third fixing element 19. As illustrated, in someembodiments, the excitation fibers 13 and the emission fibers 14,respectively, are fixed with respect to each other and with thecorresponding through-hole 45, 46 by an adhesive material 63 and may berandomly arranged. In some alternative embodiments, instead of onesecond through-hole 45 accommodating the second end portions 17 of theexcitation fibers 13 and one third through-hole 46 accommodating thesecond end portions 17 of the emission fibers 14, the second and thirdfixing elements 18, 19 can respectively be provided with onethrough-hole (not illustrated) accommodating all second end portions 17of both the excitation and emission fibers 13, 14, wherein in someembodiments, the second end portions of both the excitation and emissionfibers 13, 14 are fixed with respect to each other by an adhesivematerial. In some yet alternative embodiments, instead of one secondthrough-hole 45 accommodating the second end portions 17 of theexcitation fibers 13 and one third through-hole 46 accommodating thesecond end portions 17 of the emission fibers 14, the second and thirdfixing elements 18, 19 can respectively be provided with an array offirst and second through-holes 45, 46. Stated more particularly, thesecond fixing element 18 can be provided with an array of secondthrough-holes 45 accommodating the second end portions 17 of theexcitation fibers 13, wherein each second through-hole 45 accommodatesthe second end portion 17 of one excitation fiber 13, and the thirdfixing element 19 can be provided with an array of third through-holes46 accommodating the second end portions 17 of the emission fibers 14,wherein each third through-hole 46 accommodates the second end portion17 of one emission fiber 14. Accordingly, in the coupling arrangement 12there is a fixed one-to-one relationship or mapping between the firstend face 60 of the fixed first end portion 15 and the second end face 61of the fixed second end portion 17 of individual optical fibers 13, 14.Otherwise, in operative position of the detection module 6 allowinglight to be transmitted between the wells 4 and the optical fibers 13,14, a one-to-one relationship or mapping between the wells 4 and thesecond end faces 61 of the fibers 13, 14 is given. Due to the mapping,light can selectively be transmitted to each of the wells 4 and/orselectively received therefrom by coupling the excitation light 9 intospecific second end faces 61 of the excitation fibers 13 and couplingthe emitted light 24 out of specific second end faces 61 of the emissionfibers 14.

As illustrated in FIGS. 5 and 6 depicting enlarged detailed views of theinstrument 1, in some embodiments, the first fixing element 16 isprovided with a plate-like planar portion 72 provided with projections58 projecting towards the multi-well plate 3. In one embodiment, theprojections 58 are cylindrical in shape, and in other embodiments theprojections 58 may be rectangular or square in their cross-sectionalshape. In still other embodiments, other shapes may be used to definethe cross-sectional shape of the projections 58 so long as theprojections are sized to fit into the plate holes 67. Specifically, insome embodiments, each of the first through-holes 42 penetrates theplanar portion 72 and accommodates one projection 58 in a centeredposition. Stated more particularly, in some embodiments, one firstthrough-hole 42 accommodates the first end portions 15 of one pair ofone excitation fiber 13 and one emission fiber 14 wherein the first endfaces 60 thereof are flush with an orifice 59 of the first through-hole42. In one embodiment orifice 59 may be circular in cross-section and inother embodiments may have any other suitably shape in cross-section solong as the pair of one excitation fiber 13 and one emission fiber 14 isaccommodated therein. Moreover, in some embodiments, each of the firstthrough-holes 42 is provided with a ring-like broadened hollow 62adjacent to an upper side of the first fixing element 16 filled withadhesive material 63 in order to fix the first end portions 15 of theoptical fibers 13, 14 accommodated therein. While only one optical fiber13, 14 per first through-hole 42 is shown in FIG. 5, it is to beunderstood, that, in some embodiments, each first through-hole 42accommodates one excitation fiber 13 and one emission fiber 14 asdepicted in FIG. 6. Similarly, the second end portions 17 of the opticalfibers 13, 14 are fixed to the second and third fixing elements 18, 19,respectively, by an adhesive material which is not further detailed inthe figures. Accordingly, the optical fibers 13, 14 are not fixed inregions other than the fixed first and second end portions 15, 17. Insome alternative embodiments (not illustrated), the optical fibers 13,14 are fixed in-between the first and second end portions 15, 17, e.g.,by means of an adhesive such as, but not limited to a polyurethane foam,in order to reduce the motility of the optical fibers 13, 14.

Due to the fact that at least the first and second end portions 15, 17of the optical fibers 13, 14 are fixed with respect to each other, itcan be avoided that mechanical forces act on the optical fibers 13, 14during a vertical movement of the detection module 6. Accordingly,changes of the shape of the optical fibers 13, 14 (fiber bending) whichusually go along with undesired variations of the optical properties ofthe optical fibers 13, 14 can be avoided. Hence, the reliability andreproducibility of the detection results are improved. Otherwise,lifetime of the optical fibers 13, 14 can be prolonged.

As illustrated in FIG. 6, in some embodiments, each of the excitationand emission fibers 13, 14 is comprised of a core 76 made of opticallytransparent material such as, but not limited to fused silica or aplastic polymer. In some embodiments, the core 76 is coated by acladding (not illustrated) made of material having a lower opticalrefraction index than the core 76 so as to keep light within the core76. As illustrated, in some embodiments, the optical fibers 13, 14 areprovided with an opaque coating 77 permitting the fibers 13, 14 to bereadily stick together without removing the coating 77 facilitatingproduction of the instrument 1. In some embodiments, the cores 76 have adiameter in the range of from 0.05 mm to 1.5 mm, preferably 0.1 to 0.8mm. In some embodiments, the optical fibers 13, 14 including thecoatings 77 have a diameter in the range of from 0.3 to 2 mm, preferably0.4 mm to 1.0 mm.

Specifically, when performing the PCR, it is desirable that the samples5 have temperatures throughout the thermo-cycling process that are asuniform as reasonably possible since even small variations can cause afailure or undesirable outcome of the amplification process. Otherwise,since the wells 4 usually are not completely filled with samples 5, airgaps can be present in the wells 4 between the liquid samples 5 and thesealing cover 34. Hence, thermo-cycling can cause formation ofcondensates on the underside of the sealing cover 34 which reduces theoptical transmission of the sealing cover 34 and thus may interfere withthe optical detection of the emitted light 24. Condensates otherwise arelikely to vary the composition of the reaction mixtures.

With continued reference to FIGS. 5 and 6, in order to overcome suchdrawbacks, in some embodiments, the coupling arrangement 12 includes acover heater 64 for heating the sealing cover 34 fixed to the lowerhorizontal plates 41 of the chassis 38 in a manner to be in good thermalcontact therewith. Specifically, in some embodiments, the cover heater64 includes a heating plate 66 made of material having good thermalconductivity such as metallic material, e.g., stainless steel oraluminum. The heating plate 66 has a lower contact face 37 which inoperative position of the detection module 6 contacts the sealing cover34. In some embodiments, the heating plate 66 has a thickness in a rangeof from 2 to 7 mm. In some embodiments, the heating plate 66 has athermal conductivity of more than 5 W/m/K.

As illustrated, in some embodiments, the cover heater 64 furtherincludes a heating element 65 for generating heat attached to an upperplate face 69 of the heating plate 66. In some embodiments, the heatingelement 65 is adapted to generate Ohmic heating and, e.g., can beprovided with resistive heating lines (not illustrated).

With continued reference to FIG. 5, in some embodiments, the heatingplate 66 is provided with a plurality of plate holes 67 the number andarrangement of which correspond to the wells 4 of the multi-well plate3. As illustrated, in some embodiments, inoperative position of thedetection module 6, the contact face 37 contacts the sealing cover 34only in contact regions 71 in-between adjacent wells 4. Specifically, insome embodiments, a mechanical pressure can be exerted on the contactregions 71 by means of the contact face 37 to press the wells 4 into therecesses 33 of the receptacle 28 so as to obtain a reliable thermalcontact between the multi-well plate 3 and the thermal block 27. Ingeneral, the cover heater 64 enables minimization of temperature errorsand variations between the samples 5, in particular, by reducing edgeeffects which may cause temperature differences between outer and innerwells 4. Otherwise, formation of condensate on the sealing cover 34 canbe avoided.

In some embodiments, the controller 25 is electrically connected to thecover heater 64 by electric lines (not illustrated) to regulate adesired thermal output which, in some embodiments, is being varied inresponse to the input from one or more temperature sensors (notillustrated) for sensing the temperature of the heating plate 66. Insome embodiments, it can be preferred to regulate the thermal output ina manner that the temperature of the heating plate 66 is 5° C. to 15° C.above a maximum temperature of the reaction mixture which in case of thepolymerase chain reaction may, e.g., be in a range of from 95° C. to110° C.

With continued reference to FIGS. 5 and 6, in some embodiments, theheating element 65 is provided with a plurality of element holes 70,each of which opens into the plate holes 67 to thereby form common coverheater holes 78. As illustrated in FIG. 6, in some embodiments, theelement holes 70 are flush with the plate holes 67. In some embodiments,the cover heater holes 78 are configured as cylindrical through-holes.In some embodiments, each of the projections 58 accommodating theoptical fibers 13, 14 steps into a separate cover heater hole 78 withoutbeing in direct contact with the cover heater 64 so as to avoidconductive heat transfer to the optical fibers 13, 14.

In some embodiments, in operative condition of the detection module 6,that is to say, in a position where the contact face 37 contacts thecontact regions 71 in-between the wells 4, the cover heater holes 78form closed cavities avoiding convecting air so as to improve uniformityof the temperature of the samples 5. Otherwise, in some embodiments,plural air-filled cavities 68 are formed between a lower planar portionface 73 of the planar portion 72 of the first fixing element 16 and anupper heating element face 74 of the heating element 65. Accordingly,thermal communication between the first fixing element 16, in particularthe optical fibers 13, 14 fixed therein, and the cover heater 64 can bereduced.

In some embodiments, the cover heater holes 78 and/or the cavities 68above the cover heater 64 are at least partly filled with materialhaving poor thermal conductivity such as plastic material so as toreduce thermal coupling between the optical fibers 13, 14 and the coverheater 64. In some embodiments, the optical fibers 13, 14 are coated byor embedded in material having poor thermal conductivity such as plasticmaterial so as to reduce thermal coupling between the optical fibers 13,14 and the cover heater 64 and to prevent any even small distortion ofthe material when operating the detection module 6. In some embodiments,the material having poor thermal conductivity is used to fix the opticalfibers 13, 14 within the first through-holes 42.

The fixation of the optical fibers 13, 14 and sometimes the opticalfibers 13, 14 themselves depending on their material are sensitive toheat. Due to the fact that the optical fibers 13, 14 are largelythermally de-coupled from the cover heater 64 as above-detailed,lifetime of the optical fibers 13, 14 and their fixation can beprolonged. Similarly, thermal de-coupling of the excitation and emissionoptics 20, 21 from the cover heater 64 can also be reached. Anotherfeature is given by the fact that each well 4 optically communicateswith only one cover heater hole 78 in such a manner that the wells 4 areoptically shielded with respect to each other.

With continued reference to FIGS. 2 and 3, in some embodiments, theexcitation arrangement 7 is accommodated in a first casing 47 whichbeing fixed to an upper face of the upper horizontal plate 40, in thefollowing denoted as “upper horizontal plate face 75”, is integrallyformed with the chassis 38. The purpose of the excitation arrangement 7is to direct excitation light into the excitation fibers 13. Those ofskill in the art will appreciate that wavelength, power, homogeneity andaperture are subject to the design chosen according to the specificdemands of the user.

As illustrated in FIG. 2, in some embodiments, the excitationarrangement 7 includes two light sources 8 such as, but not limited to,light emitting diodes (LEDs) having two different wavelengths. The lightsources 8 are arranged in orthogonal relationship with respect to eachother, wherein each light source 8 is optically coupled to one or moreexcitation filters 22 for filtering the excitation light 9 incident on adichroic mirror 48. In some embodiments, the optics, e.g., is designedto generate a homogeneous illuminated spot having a diameter of 8 mm anda numerical aperture of 0.15 in order to direct light into theexcitation fibers 13. In some embodiments, two or more LEDs are coupledby dichroic mirrors and/or by fiber optics and/or by moving or rotatingelements such as moving or rotating mirrors or prisms in order to directthe light to an excitation adapter. The use of multiple colored LEDs isuseful for high excitation power. In order to measure the fluorescencewith various excitation wavelength ranges the LEDs can be switched orthe rotating elements can be rotated. In some embodiments, a white lightsource such as a halogen lamp or a white LED is used in combination witha filter wheel or filter sledge (as illustrated in FIG. 1). In someembodiments, the detection arrangement 10 is accommodated in a secondcasing 49 which being fixed to the upper horizontal plate face 75 isintegrally formed with the chassis 38. The purpose of the detectionarrangement 10 is to measure the emitted light 24 exiting the emissionfibers 14.

As illustrated, in some embodiments, the detection arrangement 10includes plural detectors 11, each of which having one light-sensitiveelement or at least one detector 11 having a plurality oflight-sensitive elements for optically detecting the emitted light 24such as, but not limited to, laterally resolving detectors like chargecoupled detectors (CCDs) and CMOS detectors, and linear array detectorswhich can be moved for scanning and two-dimensional-array sensors suchcamera sensors. The emission optics 21 is used to transmit the emittedlight 24 towards the detector 11 which, in some embodiments, isoptically coupled to one of a plurality of emission filters 23 forfiltering the emitted light 24.

With continued reference to FIG. 2, in some embodiments, plural emissionfilters 23 are attached to a filter wheel 50 which can be spun around acentral spin axis by means of electric motor 51 so as to selectivelymove one emission filter 23 into the optical path of the emitted light24. In some embodiments, the emitted light 24 is directed to multipledetectors such as CCD-cameras in parallel, e.g., after the emitted light24 has been chromatically separated by dichroic mirrors and/orconventional filters. Stated more particularly, in some embodiments, apicture of the second end faces 61 of the second end portions 17 of theemission fibers 14 is obtained by the detectors 11 without having aone-to-one mapping between the second end faces 61 and the pixels of thedetectors 11. Instead, the picture is processed to determine the lightintensity from each fiber. Based on the one-to-one mapping informationof the first and second end portions 15, 17 of the emission fibers 14,the determined light intensity can be attributed to a particular vessel.In some alternative embodiments, a plurality of detectors 11 can beassigned to the second end faces 61 of the emission fibers 14 in aone-to-one relationship so that each second end face 61 is related toone assigned detector 11.

In some embodiments, the excitation and emission optics 20, 21 includeone or more light guiding and/or light shaping and/or light directingelements (not illustrated) such as, but not limited to, lenses andplanar or bent mirrors and/or one or more light separating elements (notillustrated) such as, but not limited to, transmission gratings,reflective gratings and prisms in order to transmit the excitation light9 to the samples 5 and to detect the emitted light 24 by the pluralityof detectors 11. For this purpose, in some embodiments, the controller25 is operatively coupled to the light sources 8 and the detectors 11 tooutput control signals for emitting the excitation light 9 and detectingthe emitted light 24. Otherwise, the excitation and emission filters 22,23 can be changed according to the specific demands of the user.

With particular reference to FIGS. 7A and 7B, in some embodiments, theinstrument 1 includes an automated moving mechanism 52 allowing thedetection module 6 to be at least vertically moved relative to thethermal module 2. In some embodiments, the detection module 6 can bemoved vertically and horizontally. As illustrated, in some embodiments,the moving mechanism 52 includes two vertical guiding rails 53 which arefixed to the thermal block 27 in guiding engagement with the two lowerhorizontal plates 41 of the chassis 38 for linearly guiding thedetection module 6. In some embodiments, the moving mechanism 52 furtherincludes an actuating mechanism such as, but not limited to, a spindledrive operatively coupled to the chassis 38 for moving the detectionmodule 6 relative to the thermal module 2 along the guiding rails 53.Hence, the detection module 6 can at least vertically be moved towardsand away from the thermal module 2. As illustrated in FIG. 7A, in someembodiments, the detection module 6 can vertically be lowered intooperative position for thermally treating the samples 5 and, asillustrated in FIG. 7B, vertically raised into inoperative position inwhich the detection module 6 is distanced from the thermal module 2.Specifically, in inoperative position, a free space 54 is createdin-between the detection module 6 and the thermal module 2 allowing themulti-well plate 3 to be brought in a position on the thermal block 27for thermally treating the samples 5. In some embodiments, themulti-well plate 3 can be manually placed on the thermal block 27 orremoved therefrom. Those of skill in the art will recognize that thevertical dimensions of the free space 54 can be varied according to thespecific demands of the user.

On the other hand, in some embodiments, the moving mechanism 52 isadapted to forcibly press the detection module 6 on the thermal module2, that is to say, to apply a desired pressure force on the multi-wellplate 3. Accordingly, the wells 4 can be pressed into the recesses 33 ofthe receptacle 28 by means of the contact face 37 with a view ofimproving the thermal communication between the multi-well plate 3 andthe thermal block 27 so as to make the heat distribution uniform.Otherwise, the pressure force can improve the sealing effect of thetransparent sealing cover 34. In some embodiments, the detection module6 can be manually pressed on the multi-well plate 3. In someembodiments, the detection module 6 can be automatically pressed on themulti-well plate 3. For this purpose, in some embodiments, thecontroller 25 is electrically connected to the moving mechanism 52 tooutput control signals to regulate the automated vertical movement ofthe detection module 6. In some embodiments, the pressure force exertedon the multi-well plate 3 is in a range of from 100 N to 1000 N,preferably in a range of from 200 N to 600 N. Otherwise, in someembodiments, the detection module 6 can be manually raised ininoperative position to generate the free space 54 for the manual and/orautomated charging or uncharging of the instrument 1 with the multi-wellplate 3.

With particular reference to FIGS. 9A to 9F, in some embodiments, thethermal module 2 can be moved out in a charging/uncharging positionoutside an instrument casing 90 for charging the multi-well plate 3 onthe receptacle 28 and removing it therefrom, respectively, or in aprocessing position inside the instrument casing 90 for thermallytreating the liquid samples 5 and detecting the reaction productsobtained by means of the detection arrangement 10, e.g., provided bydetection module 6. Stated more particularly, the instrument 1 includesa horizontally movable tray 102 comprising a horizontal supporting base103 supporting the thermal module 2 and a vertical front cover 105mounted to the supporting base 103 for closing the instrument casing 90in processing position of the thermal module 2. The supporting base 103comprises one or two parallel beams 104 slidably engaged with horizontalguiding rails 113 vertically fixed to a base 89 of the instrument 1allowing the tray 102 to be slidably moved in and out of the instrumentcasing 90.

The instrument 1 further includes a bi-stable opening/closing devicegenerally referred to at reference numeral 108 for automaticallyperforming an opening or closing movement of the tray 102 and securingthe tray 102 in closed position. Specifically, the bi-stableopening/closing device 108 comprises a central turning knuckle 97 forrotatably supporting a first arm 98 and a second arm 99 around a centralaxis 109 radially projecting from the turning knuckle 97. The turningknuckle 97 is slidably supported by means of a horizontal guiding rod100 in parallel alignment with respect to the guiding rails 113 of thesupporting base 103. At their free ends, the two arms 98, 99 areinter-connected by a coil spring 101. The tray 102 can be releasablyconnected to the opening/closing device 108 by means of an elasticallydeformable spring catch 107 fixed to the supporting base 103.Specifically, the spring catch 107 forms a deepened resting portion 114for engagement with a projection 112 fixed to the opening/closing device108 so as to be movable with the turning knuckle 97. The instrument 1further includes a lever 91 fixedly secured to the instrument 1 atfulcrum 92. On the one side of the fulcrum 92, the lever 91 has an upperlever portion 93 which, at its upper end 95, is coupled to thevertically movable detector module 6 by means of connecting rod 96. Onthe other side of the fulcrum 92, the lever 91 has a lower lever portion94 which, at its lower end 110, is coupled to the horizontally movableturning knuckle 97.

With particular reference to FIG. 9A, a situation is depicted where thethermal module 2 is in processing position inside the instrument casing90 for thermally treating the samples 5. In this situation, thedetection module 6 is in the lowered operative position, e.g., applyinga force on the multi-well plate 3, for detecting the emitted light 24.The two arms of the bi-stable opening/closing device 108 are in a firststable position on the one side (in FIG. 9A, e.g., the left side) of theturning knuckle 97, in which the free ends thereof are elasticallyconnected by the coil spring 101. In the first stable position, the twoarms 98, 99 form an angle of about 90 degrees which can readily bereached by stoppers (not illustrated) for stopping the rotationalmovement of the arms 98, 99. In this situation, the tray 102 isconnected to the opening/closing device 108 by means of the spring catch107 engaged with the projection 112 which rests in the resting portion114 of the spring catch 107. Accordingly, the tray 102 is securedagainst inadvertent manual opening since the elastic force of the coilspring 101 has to be overcome when the tray 102 is to be opened.

With particular reference to FIGS. 9B and 9C, another situation isdepicted in two perspective views where the detection module 6 has beenbrought in the vertically raised inoperative position by means of thevertical moving mechanism 52. In inoperative position, the detectionmodule 6 is distanced from the thermal module 2 so as to create the freespace 54 there-between. When moving the detection module 6 in verticaldirection, the lever 91 is turned at the fulcrum 92 (in FIGS. 9B and 9C,e.g., in clockwise direction) so that the turning knuckle 97 is movedalong the guiding rod 100 (e.g. to the left side) by means of the lowerlever portion 94 coupled to the turning knuckle 97. Being coupled to theprojection 112, the tray 102 is simultaneously moved in horizontaldirection (in FIGS. 9B and 9C to the left side) so that a recessed grip106 for manually gripping the tray 102 formed by the upper side of thefront cover 105 becomes accessible from outside. Otherwise, when raisingthe detection module 6, the two arms 98, 99 are rotated beyond aninstable position in which they extend in opposite directionselastically expanding the coil spring 101.

With particular reference to FIGS. 9D and 9E, driven by the elasticforce of the contracting coil spring 101, the two arms 98, 99 arebrought in a second stable position on the other side (in FIGS. 9D and9E, e.g., the right side) of the turning knuckle 97. In the secondstable position, the free ends of the arms 98, 99 are elasticallyconnected by the coil spring 101 which again form an angle of about 90degrees which can readily be reached by stoppers for stopping rotationalmovement of the arms (not shown). As a result, the tray 102 coupled tothe opening/closing device 108 by the spring catch 107 is moved in aposition where the recessed grip 106 is fully accessible from outside.

With particular reference to FIG. 9F, by manually gripping the recessedgrip 106, the tray 102 can be drawn out of the instrument casing 90 tobe brought in the charging/uncharging position for charging themulti-well plate 3 on the receptacle 28 of the thermal module 2 orremoving it therefrom. When manually moving the tray 102 in thecharging/uncharging position, the spring catch 107 gets out ofengagement with the projection 112 by counteracting the elastic force ofthe spring catch 107 so that the tray 102 is released from the bi-stableopening/closing device 108.

Otherwise, the supporting base 103 can readily be returned into theinstrument casing 90 by the reverse action. Accordingly, the tray 102 ismanually pushed at the recessed grip 106 into the instrument casing 90until the spring catch 107 gets in engagement with the projection 112,and is further pushed to the inside so as to move the turning knuckle 97along the guiding rod 100 (e.g. to the right side) until the two arms98, 99 are rotated beyond the instable position in which they extend inopposite directions elastically expanding the coil spring 101. Nowdriven by the elastic force of the contracting coil spring 101, the twoarms 98, 99 are brought in the first stable position causing the tray102 to be automatically moved into the closed position where the frontcover 105 closes the instrument casing 90 and the thermal module 2 is inprocessing position.

In some embodiments, the wells 4 are pre-filled with the samples 5before being charged into the instrument 1. In some embodiments, thewells 4 are filled with the samples 5 when the multi-well plate 3 islocated on the receptacle 28. In some embodiments, the samples 5 are putthrough various temperature excursions to thereby incubate reactionmixtures contained therein at predefined temperatures for predefinedincubation intervals, e.g., for performing the polymerase chainreaction. The temperature of the samples 5 may, e.g., be raised toaround 90° C. for melting the nucleic acids and lowered to approximately40° C. to 70° C. for primer annealing and primer extension along thedenaturated polynucleotide strands. In some embodiments, melting of thenucleic acids is performed wherein, e.g., fluorescence light of thesamples 5 is detected while the temperature of the samples 5 is slowlyrisen or lowered. A typical melting curve may start between 30° C. and50° C. and may end between 75° C. and 95° C. wherein ramp rates in arange of from 0.05 to 0.25° C./sec can, e.g., be used.

The instrument 1 for the automated thermo-cycling of samples 5 can bemade highly-compact permitting the use of short optical fibers 13, 14.The first end faces 60 of the fixed first end portions 15 of the opticalfibers 13, 14 can be brought very close to the transparent sealing cover34 so as to improve the sensitivity of the optical detection of theemitted light 24 while avoiding any direct contact between the fibers13, 14 and the sealing cover 34. In some embodiments, a (vertical)distance between the first end faces 60 and the transparent sealingcover 34 is in the range of from 0.5 to 5 mm, preferably in the range offrom 1 mm to 3 mm.

In some embodiments, the instrument 1 can be operated to performreal-time (on-line) detection of the emitted light 24 so as to identifyreaction products of the samples 5, e.g., in parallel for all samples 5even during progress of the thermal treating. Particularly, in someembodiments, the samples 5 can be thermally cycled while synchronouslydetecting the emitted light 24 for all samples 5 in parallel. Otherwise,in some embodiments, due to the paired optical fibers 13, 14, couplingof the excitation light 9 into the excitation fibers 13 and coupling ofthe emitted light 24 out of the emission fibers 14 can be performedsynchronously and in parallel for all samples 5.

As illustrated in FIG. 10, illustrating another exemplary instrument 1,in some embodiments, both the excitation and detection arrangement 7, 10are arranged to have a vertical path of rays, wherein the excitationarrangement 7 includes only one light source 8 optically coupled to oneor more excitation filters 22 for filtering the excitation light 9.Accordingly, there is no need for using a dichroic mirror as depicted inFIG. 2 for illuminating the samples 5.

Reference is now made to FIGS. 11 and 12 to illustrate an exemplarymethod for determining a one-to-one relationship between the first andsecond end portions 15, 17 of the optical fibers 13, 14 of theinstrument 1. With particular reference to FIG. 11, in some embodiments,the first end portions 15 of the optical fibers 13, 14 are fixed withrespect to each other by the first fixing element 16 (not illustrated inFIG. 11) while the second end portions 17 of the excitation fibers 13are fixed with respect to each other by the second fixing element 18 andthe second end portions 17 of the emission fibers 14 are fixed withrespect to each other by the third fixing element 19. In FIG. 11, onetype of optical fibers 13, 14 is depicted for the purpose ofillustration only while it is to be appreciated that FIG. 11 similarlyapplies to both excitation and emission fibers 13, 14.

In some embodiments, there is a stochastic or random arrangement of thefirst and second end portions 15, 17 of the optical fibers 13, 14 ineach of the first to third fixing elements 16, 18, 19 considerablyfacilitating the production thereof. This is because no specific schemeor ordering has to be observed during the production. Hence, withrespect to the first end portions 15 of the optical fibers 13, 14 whichin operative position of the detection module 6 are in a position toestablish a one-to-one relationship between the first end portions 15and the wells 4 and, e.g., are located right above the wells 4, itinitially is not apparent into which well 4 the excitation light 9exiting the first end portions 15 of the excitation fibers 13 is beingdirected, and, typically more important, by which sample 5 the emittedlight 24 exiting specific second end portions 17 of the emission fibers14 has been generated. For instance, the emitted light 24 of a serialarrangement of eight wells 4 denoted as A, B, C, D, E, F, G and H can bedetected by a serial arrangement of second end portions 17 of theemission fibers 14 G, F, A, D, H, B, E and C. Hence, without knowing theexact mapping (one-to-one relationship) between the first end portions15 of the emission fibers 14 and the second end portions 17 thereof, thesamples 5 cannot be individually (selectively) detected. In order toovercome such drawback, the mapping between the first and second endportions 15, 17 of each of the optical fibers 13, 14 has to bedetermined.

With particular reference to FIG. 12, in some embodiments, a device 79for determining the mapping between the first and second end portions15, 17 of the optical fibers 13, 14 (in the following denoted as“mapping device”) is used. As illustrated, in some embodiments, themapping device 79 comprises a planar plate-like base 80 having an upperbase face 87 provided with a rectangular array of plural lightgenerating elements 81 such as, but not limited to diodes. In someembodiments, the number and arrangement of the light generating elements81 correspond to the wells 4 of the multi-well plate 3. In someembodiments, the light generating elements 81 are arranged in pluralparallel columns and plural parallel rows intersecting each other atright angles. As illustrated, in some embodiments, the mapping device,e.g., includes eight columns A-H and twelve rows 1-12 of lightgenerating elements 81 depending on the multi-well plate 3 used in theinstrument 1.

With continued reference to FIG. 12, in some embodiments, the mappingdevice 79 further comprises a control panel 82 and is equipped with twomanually operable turn-switches 83, 84. Specifically, in someembodiments, a first turn switch 83 can be used for setting a specificcolumn A-H and a second turn switch 84 can be used for setting aspecific row 1-12. In some embodiments, the control panel 82 is beingconnected to a device controller 85 fixed on the underside of the base80 for controlling the light generating elements 81 by means of electriclines 86. As set by the turn-switches 83, 84, the light generatingelements 81 can selectively be supplied with electric current togenerate light.

With yet continued reference to FIG. 12, in some embodiments, the base80 is configured to be placed on the receptacle 28 instead of themulti-well plate 3 in such a manner that the detection module 6 can bemoved in operative condition. In some embodiments, the first endportions 15 of the emission fibers 14 are located right above the lightgenerating elements 81 in such a manner that each first end portion 15is related to a separate light generating element 81. In someembodiments, the lower contact face 37 of the heating plate 66 contactsthe upper base face 87 in contact areas 88 in-between the lightgenerating elements 81 so that each light generating element 81 isaccommodated in a separate cover heater hole 78. Accordingly, in someembodiments, the light generating elements 81 are optically shieldedwith respect to each other incase the contact face 37 contacts the upperbase face 87.

Accordingly, knowing the exact position of each of the light generatingelements 81 as given by its column and row, in some embodiments, thelight generating elements 81 are consecutively supplied with electriccurrent to generate light that is coupled into the first end portions 15of the emission fibers 14 and coupled out at the second end portions 17thereof. Accordingly, by means of the detectors 11 detecting the lightof the light generating elements 81, a one-to-one relationship (mapping)between the first end portions 15 of the emission fibers 14 or wells 4and the second end portions 17 can readily be established. In someembodiments, this one-to-one relationship is saved in a, e.g. permanent,data storage 115, e.g., in the form of a look-up table so as to enablethe wells 4 or samples 5 contained therein to be selective detected.

As above-detailed, in some embodiments, a picture of the second endfaces 61 of the second end portions 17 of the emission fibers 14 isobtained by the detectors 11 without having a one-to-one mapping betweenthe second end faces 61 and the pixels of the detectors 11. Otherwise,electronic picture processing, e.g., based on the known one-to-onerelationship (mapping) between the first and second end portions 15, 17of the emission fibers 14 can be used to obtain information about therelationship between the pixels of the detectors 11 and the second endfaces 61 of the emission fibers 14 so as to attribute the detected lightto individual wells 4.

With particular reference to FIGS. 13 to 15, a clamping mechanism 129for clamping the receptacle 28 of the instruments 1 of FIGS. 2 to 10 isdescribed. As a matter of fact, in some embodiments such as, but notlimited to, the case of using the instrument 1 as thermo-cycler, thereceptacle 28 is required to have a temperature distribution ashomogeneous as reasonably possible. In order to meet this requirement,the receptacle 28 is clamped to the one or more thermoelectric devices55 of the thermal block 27. The clamping mechanism 129 is configured toexert a well-defined force and force distribution to the receptacle 28.Stated more particularly, in some embodiments, the clamping mechanism129 comprises a resilient clamp 123 made of elastic material such as,but not limited to, spring steel according to DIN regulation 1.4310. Insome embodiments, the clamp 123 is configured as flat spring.Specifically, as illustrated, in some embodiments, the clamp 123 isconfigured as elongate member having two opposing arms 126 connected bya middle portion 127. The middle portion 127 is provided with anisolation block 124 resting on a base plate 125 of the receptacle 28.The isolation block 124 is made of thermally isolating material such as,but not limited to, fiber enforced polymer. Flanges 119 on both sides ofthe clamp 123 are provided with gripping recesses 130 in grippingengagement with gripping portions 128 of the arms 126. The flanges 119are fixedly secured to the thermal block 27 by means of screws 120 whichcan be screwed in and out of mounting holes 121. Accordingly, by turningthe screws 120 in, the gripping portions 128 of the arms 126 can bemoved towards the base plate 125 while bending the arms 125. As aresult, a clamping force is exerted on the base plate 125 via the middleportion 127 to thereby clamp the receptacle 28 to the one or morethermoelectric devices 55 to thereby equalize mechanical tolerances. Asillustrated, in some embodiments, a further homogenization of theclamping force and thus temperature distribution can be reached by oneor more interface layers 122 on one or both sides of the thermoelectricdevices 55 made of material having good thermal conductivity such as,but not limited to, oil, paste or foil adapted for heat transfer. Thethermal isolation block 124 inhibits excessive heat-flow from thereceptacle 28 towards the thermal block 27 by means of the clamp 123,e.g., in a case when the receptacle 28 is heated to a temperature of,e.g., 95° C. and the thermal block 27 has a lower temperature of, e.g.,35° C. In some embodiments, e.g., for a thermoelectric device 55 havingan area of about 16 cm², the clamp 123 is configured to exert a clampingforce in a range of from 200 N to 500 N. Those of skill in the art,however, will appreciate that the clamping force exerted on thereceptacle 28 may vary according to the specific demands of the userand, in some embodiments, can for instance be in a range of from 10 N to1000 N. In some embodiments, instead of one clamping mechanism 129, theinstrument 1 includes plural clamping mechanisms 129 for clamping thereceptacle 28 to the thermoelectric devices 55.

Obviously many modifications and variations of the present invention arepossible in light of the above description. It is therefore to beunderstood, that within the scope of appended claims, the invention maybe practiced otherwise than as specifically devised. Some examples: insome embodiments, the excitation and detection arrangements 7, 10 arenot integrally formed with the coupling arrangement 12 permitting thelight-weight coupling arrangement 12 to be vertically moved intooperative position for treating the samples 5 or inoperative positionfor charging/uncharging the multi-well plate 3 while keeping theexcitation and detection arrangements 7, 10 unmoved. This requires thesecond end faces 61 of the fibers 13, 14 to be optically coupled to theexcitation and detection arrangements 7, 10. In some embodiments, thedetection module 6 is laterally arranged as to the thermal module 2 sothat the constructional height can be reduced. In some embodiments,instead of a pair of optical fibers 13, 14 per well 4, only one opticalfiber is used for transmitting both the excitation and emission light.In these cases, the excitation and emitted light can be opticallyde-coupled by means of a dichroic mirror. In some embodiments, a bundleof optical fibers 13, 14 per well 4 is used for transmitting theexcitation and emitted light 9, 24. In some embodiments, instead ofbeing perpendicularly arranged with respect to the wells 4, the firstthrough-holes 42 are inclined with respect to a (e.g. horizontal)opening face of the wells 4. In some embodiments, instead ofaccommodating one pair of optical fibers 13, 14 in one through-hole 42,45, 46, each fiber 13, 14 is accommodated in a separate through-hole sothat each pair of excitation and emission fibers 13, 14 is accommodatedin two through-holes. In these cases, the through-holes of one pair ofoptical fibers 13, 14 may, e.g., have a minimum distance of 0.2 mm. Thislist of modified embodiments is not exhaustive.

What is claimed is:
 1. An instrument for the automated thermal treatmentof liquid samples, comprising: a temperature-controlled receptacle forloading with a plurality of vessels for containing said samples, saidreceptacle being configured to form a thermal communication with saidloaded vessels; a detection module equipped with a detection arrangementprovided with one or more detectors for detecting light emitted fromsaid samples and a coupling arrangement provided with a plurality ofoptical fibers for transmitting said emitted light to said detectionarrangement, said optical fibers including emission fibers fortransmitting said emitted light to said detection arrangement andexcitation fibers for transmitting excitation light to said sampleswherein said optical fibers have first and second end portions, saidfirst end portion and said second end portion of each optical fiberbeing fixed with respect to each other, said first end portions of saidexcitation and emission fibers are fixed by at least one plate-likefixing element comprising a planar portion with projections projectingtoward the plurality of vessels and said second end portions of saidexcitation and emission fibers being fixed by at least one otherplate-like fixing element, the at least one plate-like fixing elementand at least one other plate-like fixing element being fixed relative toeach other; and a moving mechanism for moving at least one of saidcoupling arrangement and said receptacle in a manner to vary aninter-distance between said coupling arrangement and said receptacle soas to allow the vessels to be loaded to or unloaded from said receptacleand to allow detection of light from samples contained in said one ormore receptacle-loaded vessels; wherein the second end portions have astochastic lateral position and wherein a mapping between said first endportion and said second end portion of individual optical fibers isbeing stored in a data storage.
 2. The instrument according to claim 1,wherein said moving mechanism is configured to move said couplingarrangement while said receptacle is being kept stationary.
 3. Theinstrument according to claim 2, wherein said moving mechanism isconfigured to move said detection module.
 4. The instrument according toclaim 2, wherein said optical fibers include emission fibers fortransmitting said emitted light to said detection arrangement andexcitation fibers for transmitting excitation light to said samples. 5.The instrument according to claim 3, wherein said optical fibers includeemission fibers for transmitting said emitted light to said detectionarrangement and excitation fibers for transmitting excitation light tosaid samples.
 6. The instrument according to claim 1, wherein saidcoupling arrangement is being equipped with a cover heater for heating asealing cover placed over a multi-well plate providing the plurality ofvessels for containing said samples.
 7. The instrument according toclaim 1, wherein said coupling arrangement is being equipped with acover heater for heating a sealing cover placed over a multi-well plateproviding the plurality of vessels for containing said samples.
 8. Theinstrument according to claim 7, wherein said cover heater includes aheated plate-like heating member configured to be brought in physicalcontact with said sealing cover, said heating member being equipped witha plurality of openings accommodating first end portions of said opticalfibers.
 9. The instrument according to claim 8, wherein said opticalfibers are being thermally isolated from said heating member.
 10. Theinstrument according to claim 9, wherein said openings are configured toform cavities in case said heating member contacts said multi-wellplate, said cavities being adapted to optically shield said plurality ofvessels from each other.
 11. The instrument according to claim 9,wherein said heating member is configured to exert mechanical pressureon said multi-well plate so as to press said plurality of vessels intorecesses of said receptacle.
 12. The instrument according to claim 8,wherein said openings are configured to form cavities in case saidheating member contacts said multi-well plate, said cavities beingadapted to optically shield said plurality of vessels from each other.13. The instrument according to claim 8, wherein said heating member isconfigured to exert mechanical pressure on said multi-well plate so asto press said plurality of vessels into recesses of said receptacle. 14.The instrument according to claim 1, wherein said coupling arrangementis being provided with said data storage device.